One major goal of this proposal is to understand how catecholamine-sensitive adenylate cyclase systems function at the molecular level. The stoichiometry of the known components of a functioning system (beta-receptor, cyclase, guanine nucleotide regulatory site(s)) will be quantitatively determined by direct identification of each of these components following detergent solubilization and chromatographic isolation of hormone- and/or guanine nucleotide-promoted complexes of the individual components. The role of this stoichiometry in defining the extent and characteristics of nucleotide and hormone responsiveness as well as the pharmacological specificity of a system will be evaluated in systems where the extent of receptor-cyclase coupling changes markedly, e.g. during maturation of rat reticulocytes to mature erythrocytes. A second major goal is to determine whether muscarinic cholinergic attenuation of catecholamine stimulated effects in the myocardium results from perturbation of beta-receptor-nucleotide site complex formation and/or direct interaction of the muscarinic receptor with the nucleotide site. Furthermore, the possible direct interaction of beta-adrenergic and muscarinic receptors in cardiac membranes will be explored using crosslinking reagents as rulers of inter-molecular distance. Thus these studies should provide insights into the molecular basis of the regulation of myocardial function by adrenergic and cholinergic agents. Elucidation of the molecular events responsible for beta-receptor-cyclase "coupling" and the modulation of this coupling by muscarinic agents will provide a well characterized model of the way in which all hormone receptors coupled either to adenylate cyclase activation or attenuation of cyclase stimulation operate at the biochemical level. Also, these studies will identify at least one mechanism for transfer of information across the biological membrane.